ANOVA for parents and backcross progenies
In Tables 2 and 3, the ANOVA for the parents and BC2F1 progenies for 20 plant growth, yield, fruit, seed and biochemical characteristics is provided, respectively. Indicating that there is enough innate variation among the parents that it may be tapped by selection, the ANOVA showed greater significant treatment mean squares (MS) attributable to parents for all the examined traits. The treatment mean squares attributed to BC2F1 populations showed non-significant results for the number of primary branches, significant results at the 5% level for plant spread and fruit pericarp thickness, and significant results at the 1% level of significance for the remaining traits studied, illustrating the different responses of various BC2F1 populations to the studied traits. This concurs with the findings of Usman et al. (2014) and Santosh (2013).
Table 2
Analysis of variance for plant growth, yield and quality traits of parental lines
Source of variation | df | Plant height (cm) (PH) | Plant spread (cm) (PW) | Number of primary branches (NB) | Number of fruits plant− 1 (FPP) | Fruit length (cm) (FL) | Fruit width (cm) (FW) | Fruit pericarp thickness (mm) (FPT) | Ten fruit weight (g) (TFW) | Fruit yield plant− 1 (g) (FYP) | Number of locules plant− 1 (NC) |
Replication | 2 | 25.34 | 39.69 | 0.14 | 5.57 | 0.16 | 0.01 | 0.01 | 1929.09 | 9423.25 | 0.14 |
Genotypes | 6 | 4980.34** | 219.11** | 1.08** | 319.41** | 1.63** | 10.26** | 3.06** | 280839.43** | 515266.25** | 1.87** |
Error | 12 | 37.5 | 11.99 | 0.19 | 7.29 | 0.11 | 0.12 | 0.16 | 1211.73 | 13559.58 | 0.25 |
Table 2
Source of variation | df | Dry matter content in green fruits (%) (DMG) | Dry matter content in red fruits (%) (DMR) | Vitamin C content (mg 100g− 1) (VC) | Capsaicin content (%) (CC) | Chlorophyll a (mg 100g− 1) (CHA) | Chlorophyll b (mg 100g− 1) (CHB) | Total chlorophyll (mg 100g− 1) (CHT) | 1000 seed fresh weight (g) (SFW) | 1000 seed dry weight (g) (SDW) | Number of seeds fruit− 1 (SPF) |
Replication | 2 | 0.15 | 0.06 | 29.054 | 0.0002 | 0.005 | 0.02 | 0.002 | 0.04 | 0.003 | 31 |
Genotypes | 6 | 65.19** | 63.93** | 183.19** | 0.07** | 0.31** | 1.19** | 2.56** | 25.83** | 10.09** | 197.56** |
Error | 12 | 0.29 | 0.27 | 5.62 | 0.0002 | 0.002 | 0.02 | 0.03 | 0.07 | 0.05 | 17.06 |
Note: Data is mean sums of square; df: Degree of freedom; *significant at p = 0.05; **significant at p = 0.01 |
Table 3
Analysis of variance for plant growth, yield and quality traits of BC2F1 population of bell pepper
Source of variation | df | Plant height (cm) (PH) | Plant spread (cm) (PW) | Number of primary branches (NB) | Number of fruits plant− 1 (FPP) | Fruit length (cm) (FL) | Fruit width (cm) (FW) | Fruit pericarp thickness (mm) (FPT) | Ten fruit weight (g) (TFW) | Fruit yield plant− 1 (g) (FYP) | Number of locules plant− 1 (NC) |
Replication | 2 | 18.23 | 8.04 | 0.39 | 8.17 | 0.03 | 0.53 | 0.02 | 2107.31 | 8059.64 | 0.39 |
Genotypes | 5 | 1096.75** | 51.03* | 0.36 | 135.03** | 1.55** | 3.04** | 0.85* | 125495.25** | 178302.85** | 0.99** |
Error | 10 | 24.75 | 9.59 | 0.19 | 4.5 | 0.24 | 0.19 | 0.23 | 820.81 | 178302.8 | 0.13 |
Table 3
Source of variation | df | Dry matter content in green fruits (%) (DMG) | Dry matter content in red fruits (%) (DMR) | Vitamin C content (mg 100g− 1) (VC) | Capsaicin content (%) (CC) | Chlorophyll a (mg 100g− 1) (CHA) | Chlorophyll b (mg 100g− 1) (CHB) | Total chlorophyll (mg 100g− 1) (CHT) | 1000 seed fresh weight (g) (SFW) | 1000 seed dry weight (g) (SDW) | Number of seeds fruit− 1 (SPF) |
Replication | 2 | 0.06 | 0.07 | 2.84 | 0.000004 | 0.002 | 0.008 | 0.012 | 0.07 | 0.01 | 14 |
Genotypes | 5 | 2.37** | 26.41** | 53.18** | 0.00008** | 0.23** | 0.56** | 1.39** | 18.32** | 6.79** | 211.07** |
Error | 10 | 0.28 | 0.17 | 2.53 | 0.000005 | 0.002 | 0.006 | 1.39 | 0.19 | 0.03 | 20.07 |
Note: Data is mean sums of square; df: Degree of freedom; *significant at p = 0.05; **significant at p = 0.01 |
Assessment of genetic variability parameters |
The extent of crop improvement depends on the variability exists in the base plant material. In the absence of which there will be no selection response, and ultimately no crop improvement. The fruit yield is the crucial factor in the improvement of bell pepper crop and it is the outcome of many of the studied traits, hence variability for the component traits is very critical in addition to yield. Here we evaluated the six BC2F1 populations along with seven parental lines for 20 different important traits and their genetic variability measured in terms of genotypic and phenotypic variances, coefficient of genetic and phenotypic variability (GCV & PCV, respectively), broad sense heritability (H2), genetic advance and genetic gain (Tables 4 and 5).
Genotypic and phenotypic variances
The lowest and highest genotypic variance (GV, σ2g) and phenotypic variance (PV, σ2р) for parental lines was observed for capsaicin content (0.023 & 0.023, respectively) and fruit yield plant− 1 (167235.6 & 180795.1, respectively). The similar trend was observed in BC2F1 populations where GV and PV was lowest for capsaicin content (zero for both) and highest for fruit yield plant− 1 (566632.24 & 65038.37, respectively). Besides from fruit yield plant− 1 the highest GV and PV values in parental population was observed for ten fruit weight followed by plant height, number of fruits plant− 1, plant spread, seeds fruit− 1, vitamin C content and dry matter content in green and red fruits (Table 4). Likewise, the highest GV and PV values in BC2F1 populations was recorded for ten fruit weight followed by plant height, seeds fruit− 1 and number of fruits plant− 1. The moderate extent of GV and PV values were seen for vitamin C content followed by plant spread, dry matter in green fruits and 1000 seed fresh weight (Table 5). However, the lowest GV and PV values were noted for most of the studied quality attributes both in parental lines and backcross populations. Significant GV values indicate a greater genetic component to the overall variance; therefore those traits would be taken into consideration and used for selection. On the contrary, high PV suggests that environmental variables had a significant impact on the expression of the traits during the development and reproductive phases Sran and Jindal (2019).
Table 4
Estimation of heritability, genotypic, and phenotypic coefficients of variance, and genetic advance for 20 traits in bell pepper parents
Traits* | GV | GCV (%) | PV | PCV (%) | h²bs | GA 5% | Genetic gain (5%) |
PH (cm) | 1647.611 | 41.139 | 1685.111 | 41.605 | 0.978 | 82.681 | 83.799 |
PW (cm) | 69.039 | 16.213 | 81.032 | 17.565 | 0.852 | 15.799 | 30.829 |
NB | 0.294 | 21.074 | 0.492 | 27.279 | 0.597 | 0.862 | 33.536 |
FPP | 104.04 | 47.285 | 111.333 | 48.914 | 0.934 | 20.312 | 94.162 |
FL (cm) | 0.506 | 10.359 | 0.622 | 11.48 | 0.814 | 1.323 | 19.256 |
FW (cm) | 3.378 | 39.265 | 3.5 | 39.967 | 0.965 | 3.72 | 79.466 |
FPT (mm) | 0.969 | 28.788 | 1.125 | 31.022 | 0.861 | 1.882 | 55.033 |
TFW (g) | 93209.23 | 57.434 | 94420.96 | 57.806 | 0.987 | 624.873 | 117.553 |
FYP (g) | 167235.6 | 46.871 | 180795.1 | 48.734 | 0.925 | 810.219 | 92.863 |
NC | 0.54 | 23.375 | 0.794 | 28.346 | 0.68 | 1.248 | 39.707 |
DMG (%) | 21.631 | 75.766 | 21.928 | 76.284 | 0.986 | 9.516 | 155.018 |
DMR (%) | 21.222 | 38.184 | 21.49 | 38.424 | 0.988 | 9.431 | 78.167 |
VC (mg/100g) g | 59.194 | 12.241 | 64.811 | 12.808 | 0.913 | 15.147 | 24.099 |
CC (%) | 0.023 | 165.884 | 0.023 | 166.669 | 0.991 | 0.312 | 340.113 |
CHA (mg/100g) | 0.104 | 53.713 | 0.106 | 54.231 | 0.981 | 0.658 | 109.593 |
CHB (mg/100g) | 0.387 | 74.829 | 0.417 | 77.667 | 0.928 | 1.235 | 148.516 |
CHT (mg/100g) | 0.844 | 64.174 | 0.871 | 65.21 | 0.968 | 1.862 | 130.097 |
SFW (g) | 8.586 | 31.712 | 8.659 | 31.847 | 0.992 | 6.011 | 65.051 |
SDW (g) | 3.348 | 26.866 | 3.403 | 27.085 | 0.984 | 3.739 | 54.897 |
SPF | 60.167 | 12.312 | 77.222 | 13.949 | 0.779 | 14.104 | 22.388 |
h²bs: broad sense heritability, GA: Genetic advance, * for traits acronyms see Table 2 & 3 |
Genotypic and phenotypic coefficient of variances
All of the characteristics under consideration in the current study showed larger PCV values than GCV values (Tables 4 and 5), which is likely a result of environmental factors that may have an impact on how traits are expressed phenotypically. Among the parents GCV values was ranged from 10.36 to 165.88, similarly PCV values varied from 11.44 to 166.67 for fruit length and capsaicin content, respectively. The highest GCV and PCV value of capsaicin content is due to the inclusion of hot and bell type peppers in the current experiment. Values of GCV and PCV are classified as high if they are higher than 20%, medium if they are between 10% and 20%, and low if they are less than 10% (Deshmukh et al. 1986). According to this, the perusal of data from Table 4 indicates that most of the characters under investigation for parental lines have high GCV and PCV values except for plant spread, fruit length, vitamin C content, and seeds fruit− 1, demonstrating moderate GCV and PCV values. While incase of BC2F1 populations GCV and PCV values found lowest for vitamin C content (6.02 and 6.46, respectively) and highest for chlorophyll b (45.97 and 46.73, respectively). Rani (2020) also had similar study that reported highest GCV and PCV for chlorophyll b among the other studied traits. High values of GCV and PCV were an indication of high genetic variability among the developed BC2F1 populations thus there is scope for improvement of these characters through simple selection (Ishaya et al. 2020). However, in backcross populations the lowest GCV values were depicted by plant spread, number of primary branches and fruit length. The moderate amount of GCV and PCV were observed for fruit pericarp thickness, dry matter in green fruits, capsaicin content and seeds fruit− 1 (Table 5).
Heritability (H2)
The genotypic coefficient of variation measures the total genetic variance, whereas heritability assesses the degree to which a trait's genetic variability is passed down to progeny. Heritability is therefore a criterion that breeders pay close attention to since its magnitude reveals how accurately a genotype will be classified based on its phenotypic traits. Very high heritability is defined as being over 80%, high heritability as being between 60 and 79%, and medium heritability as being between 40 and 59% (Singh 2001). The review of data from Table 4 and 5 reveals that heritability estimates ranged from 59.7% (number of primary branches) to 99.2% (1000 seed fresh weight) for parental lines likewise for BC2F1 populations it was varied from 22.7% (number of primary branches) to 98.4% (1000 seed dry weight). According to the scale given by Singh (2001) most of the studied fruit and yield traits along with quality traits demonstrated very high heritability, indicating that these traits might be controlled by additive gene action. Hence, it proves that the larger proportion of phenotypic variance has been attributed to genotypic variance and it was clear that reliable selection was made for almost all the important fruit, quality and yield related traits on the basis of phenotypic expression. For instance, the negative selection for reduced capsaicin content was proved to be fruitful in achieving low pungency in advanced backcross populations. Such results are line with the findings of Mishra et al. (2005) and Sharma et al. (2019) where they reported high PCV and GCV values coupled with high heritability for fruit and yield related characters. On the contrary, medium and low heritability was recorded for number of primary branches incase of parents and backcross populations, respectively. The selection would be virtually impractical or considerably difficult for a character coupled with low heritability because of masking effect of environment on the genotypic effects.
Genetic advance (GA)
Unaccompanied heritability provides no hint of the degree of genetic change that will arise from choosing individual genotypes. Consciousness of genetic development combined with heritability is thus most useful (Lush 1949; Jhonson et al. 1955). Estimates of genetic advance among the parental lines (Table 4) for 20 studied traits exhibited a wide range of values varied from 0.312 for capsaicin content to 810.22 for fruit yield plant− 1 and in BC2F1 population (Table 5) it was ranged from 0.009 for capsaicin content to 457.45 fruit yield plant− 1. Johnson et al. (1955) classified the estimate of genetic advance as low when it was below 10%, moderate when it was from 10 to 20% and high when it was above 20%. Hence, from the present investigation it is demonstrated that high genetic advance was depicted by number of fruits plant− 1 (only for parental lines), plant height, ten fruit weight and fruit yield plant− 1 in both parental and backcross populations. In case of parental lines the moderate genetic advance values was recorded for plant spread, vitamin C content and seeds fruit− 1, similarly it was true for number of fruits plant− 1 and seeds fruit− 1 in BC2F1 populations. While, all other traits
Table 5
Estimation of genetic parameters for 20 horticultural traits in BC2F1 population of bell pepper
Traits | GV | GCV (%) | PV | PCV (%) | h²bs | GA 5% | Genetic gain (5%) |
PH (cm) | 357.334 | 23.917 | 382.084 | 24.732 | 0.935 | 37.658 | 47.647 |
PW (cm) | 13.814 | 7.784 | 23.403 | 10.131 | 0.59 | 5.883 | 12.319 |
NB | 0.056 | 9.223 | 0.244 | 19.347 | 0.227 | 0.231 | 9.058 |
FPP | 43.511 | 38.425 | 48.011 | 40.363 | 0.906 | 12.936 | 75.355 |
FL (cm) | 0.436 | 9.714 | 0.679 | 12.122 | 0.642 | 1.09 | 16.035 |
FW (cm) | 0.951 | 20.094 | 1.137 | 21.974 | 0.836 | 1.837 | 37.853 |
FPT (mm) | 0.207 | 12.144 | 0.437 | 17.627 | 0.475 | 0.646 | 17.234 |
TFW (g) | 41558.14 | 35.381 | 42378.96 | 35.729 | 0.981 | 415.861 | 72.176 |
FYP (g) | 56632.24 | 26.422 | 65038.37 | 28.315 | 0.871 | 457.453 | 50.79 |
NC | 0.289 | 17.59 | 0.411 | 20.984 | 0.703 | 0.928 | 30.376 |
DMG (%) | 0.695 | 14.221 | 0.979 | 16.875 | 0.71 | 1.448 | 24.687 |
DMR (%) | 8.746 | 26.336 | 8.915 | 26.59 | 0.981 | 6.034 | 53.735 |
VC (mg/100g) g | 16.884 | 6.023 | 19.411 | 6.458 | 0.87 | 7.894 | 11.572 |
CC (%) | 0 | 16.701 | 0 | 18.301 | 0.833 | 0.009 | 31.397 |
CHA (mg/100g) | 0.075 | 38.328 | 0.077 | 38.939 | 0.969 | 0.555 | 77.716 |
CHB (mg/100g) | 0.183 | 45.966 | 0.189 | 46.733 | 0.967 | 0.867 | 93.135 |
CHT (mg/100g) | 0.459 | 41.202 | 0.468 | 41.578 | 0.982 | 1.384 | 84.107 |
SFW (g) | 6.046 | 26.104 | 6.232 | 26.502 | 0.97 | 4.989 | 52.965 |
SDW (g) | 2.251 | 23.001 | 2.288 | 23.191 | 0.984 | 3.065 | 46.994 |
SPF | 63.667 | 12.533 | 83.733 | 14.373 | 0.76 | 14.333 | 22.512 |
h²bs: broad sense heritability, GA: Genetic advance, * for traits acronyms see Table 2 |
exhibited low genetic advance for both parental and backcross populations. It is more accurate and dependable to anticipate the outcomes and effects of selection when heritability and genetic advance are combined. Accordingly, in present investigation plant height, ten fruit weight and fruit yield plant− 1 depicted high heritability and high estimates of genetic advance in both parental lines and BC2F1 populations which suggests that these traits are under the additive gene effects. Thus, above mentioned characters could be improved through selection. Several other workers also received high heritability and high genetic advance for total fruit yield plant− 1 (Bharadwaj et al. 2007), for fruit weight, for number of fruits plant− 1 (Sreelathakumary and Rajamony 2002, 2004). On the other hand, high heritability coupled with low genetic advance was noted for fruit length, number of locules fruit− 1, dry matter content in green fruits, capsaicin content, chlorophyll a & b, and 1000 seed fresh and dry weight in both parental and backcross populations. Hence, these traits were under the control of non-additive gene actions; therefore mentioned traits could be improved through hybridization followed by selection (Ara et al. 2009).
Genetic gain
Although assessments of high heritability are useful in determining the basis for genes that are passed down from parental lines to offspring, accurate results are obtained when heredity and genetic gain are taken into account. The data given in Table 4 indicated that all the studied traits in parental lines were exhibited high genetic gain except for fruit length (moderate genetic gain) and its values varied from 19.26 (fruit length) to 340.11 (capsaicin content). However, data for BC2F1 populations in Table 5 indicated that plant spread, fruit length, and vitamin C content were revealed moderate genetic gain and number of primary branches showed low genetic gain, while other characters in BC2F1 population exhibited high genetic gain. In BC2F1 populations, the moderate heritability and moderate genetic advance was found for plant spread, fruit length and fruit pericarp thickness, while low heritability coupled with low genetic advance was only seen for number of primary branches. This showed that the non-additive gene effects are strongly influenced the certain traits. Improvement for these traits can be accomplished by further partitioning the genetic variation and allowing selection in segregating generations for the appropriate types.
High heritability coupled with high values of GCV and PCV along with high estimates of genetic advance and high genetic gain was noted for plant height, number of fruits plant− 1, fruit width, ten fruit weight, fruit yield plant− 1, number of locules fruit− 1, dry matter in green, and red fruits, capsaicin content, chlorophyll a, b and total chlorophyll, 1000 seed dry, and fresh weight in both parental lines and BC2F1 populations. It suggests that these attributes were strongly influenced by additive gene action; therefore straightforward selection based on the phenotypic expression of these traits would be preferable. Additionally, it proposed that a high-yielding LCVD bell pepper line might be chosen from the created backcross series (BC2F1).
Recovery of recurring parent characteristics in BC2F1 progenies
By comparing the horticultural and morphological performances of the improved lines carrying the LCVD resistant gene along with the adaptable recurrent parent, paired "t" test analysis was used to confirm the successful revival of the recurrent parent genome (RPG) in the BC2F1 populations. The results are provided in Tables 6, 7, 8 and 9.
Assessment of BC2F1 populations for plant growth traits
Plant height is regarded as a key yield-attributing characteristic because it encourages the growth of additional branches and prolongs the harvest period, both of which increase productivity. Compared to dwarf plants, taller plants are less likely to contract diseases. In terms of plant height, the six recipient parental lines had an average performance ranging from 51.67 cm (PAU SM-58) to 114.33 cm (PAU SM-17). Correspondingly, BC2F1 plants ranged in height from 47.73 cm (LCVRS3SM-58) to 103.75 cm (LCVRS3SM-17), this was also reported by Rani (2020). The LCVRS3SM-17 population exhibited a maximum height that was followed by LCVRS3SM-8 (92.08 cm), while the shortest plants were recorded for LCVRS3SM-58 followed by LCVRS3SM-4 (71.42 cm). Furthermore, plant spread for parental genotypes varied from 44.33 cm to 65.90 cm. The maximum plant spread was noted for PAU SM-8 followed by PAU SM-2 (54.17 cm), in contrast minimum plant spread was recorded for PAU SM-17 followed by PAU SM-1 (50.83 cm). In case of BC2F1 populations plant spread was lie between 42.56 cm (LCVRS3SM-17) and 55 cm (LCVRS3SM-8). Besides, number of primary branches plant− 1 is also important plant growth trait that
Table 6
Recovery of recurrent parent attributes in BC2F1 population of bell pepper for plant growth traits (appraise similarity through paired t-test)
Pairs | Individuals | PH (cm) | PW (cm) | NB |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 85.67 | 3.18 | 3.12ns | 0.09 | 95.33 | 50.83 | 1.45 | 0.77ns | 0.52 | 96.40 | 2.67 | 0.33 | 1ns | 0.42 | 87.27 |
LCVRS3SM-1 (BC2F1) | 81.67 | 4.17 | 49.00 | 1.15 | 2.33 | 0.33 |
2 | PAU SM-2 | 78.67 | 5.24 | 0.18ns | 0.87 | 98.60 | 54.17 | 2.05 | 6.61* | 0.02 | 86.12 | 2.00 | 0.00 | -1ns | 0.42 | NA |
LCVRS3SM-2 (BC2F1) | 77.57 | 1.62 | 46.65 | 2.09 | 2.33 | 0.33 |
3 | PAU SM-8 | 97.67 | 3.71 | 8.54* | 0.01 | 94.28 | 65.90 | 1.93 | 7.51* | 0.02 | 83.46 | 2.67 | 0.33 | -1ns | 0.42 | NA |
LCVRS3SM-8 (BC2F1) | 92.08 | 3.09 | 55.00 | 1.56 | 3.00 | 0.00 |
4 | PAU SM-4 | 82.33 | 1.86 | 11.36** | 0.01 | 86.75 | 53.33 | 2.20 | 3.08ns | 0.09 | 88.24 | 3.33 | 0.33 | 1ns | 0.42 | 90.09 |
LCVRS3SM-4 (BC2F1) | 71.42 | 1.83 | 47.06 | 0.71 | 3.00 | 0.00 |
5 | PAU SM-17 | 114.33 | 1.76 | 2.27ns | 0.15 | 90.75 | 44.33 | 1.09 | 1.35ns | 0.31 | 96.01 | 2.00 | 0.00 | -1ns | 0.42 | NA |
LCVRS3SM-17 (BC2F1) | 103.75 | 3.32 | 42.56 | 0.96 | 2.33 | 0.33 |
6 | PAU SM-58 | 51.67 | 0.88 | 2.04ns | 0.18 | 92.37 | 51.67 | 3.94 | 0.8ns | 0.51 | 89.47 | 2.00 | 0.00 | -1ns | 0.42 | NA |
LCVRS3SM-58 (BC2F1) | 47.73 | 1.83 | 46.23 | 3.02 | 2.33 | 0.33 |
Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
has direct effect on yield. Number of primary branches plant− 1 of parental lines was almost same and ranged between 2 (PAU SM-2, PAU SM-17 and PAU SM-58) to 3.33 (PAU SM-4). Among the six developed BC2F1 populations 2.33 is average number of primary branches plant− 1 except for LCVRS3SM-4 and LCVRS3SM-8. The paired t test (Table 6) revealed non-significant differences between mean values of recurrent parents and developed backcross populations for plant growth traits except for LCVRS3SM-8 and LCVRS3SM-4 for plant height and LCVRS3SM-2 and LCVRS3SM-8 for plant spread. Therefore, the results were confirmed that the performance of backcrossed plants was recovered when compared to their respective recurrent parents.
Assessment of BC2F1 populations for fruit and yield attributes
One of the most crucial characteristics that directly affect fruit yield plant− 1 is fruit weight. Medium-sized fruits (50–80 g) are often preferable in India's marketing system. Hence development of high yielding genotypes along with good fruit characteristics is help to boost the farmers’ income. The developed BC2F1 populations per se performance for ten fruit weight was ranged from 310.55 g (LCVRS3SM-8) to 804.91 g (LCVRS3SM-58), and in the case of the parents, a similar pattern was observed in which the maximum ten fruit weight was demonstrated by PAU SM-58 (861.83 g), while the minimum ten fruit depicted by PAU SM-8 (340.50 g) (Table 7). In terms of fruit length, the backcross generations LCVRS3SM-4 (5.96 cm) and LCVRS3SM-1 (7.51 cm) in BC2F1 populations recorded the lowest and highest mean values, whereas PAU SM-4 (5.58 cm) and PAU SM-2 (7.64 cm) in parental lines recorded the minimum and maximum fruit lengths, respectively. Parental lines' per se fruit width performance spanned 3.42 cm (PAU SM-8) to 6.66 cm (PAU SM-58), similar trend was observed for BC2F1 populations performance where fruit width was ranged from 3.28 cm (LCVRS3SM-8) to 5.85 cm (LCVRS3SM-58). The similar findings regarding fruit width was also reported by Rani (2020). Furthermore, fruit pericarp thickness of recurrent parents ranged from 3.47 mm in PAU SM-8 to 4.13 mm in the PAU SM-1, which of the backcrossed population ranged from 3.21 mm in LCVRS3SM-2 to 4.70 mm in LCVRS3SM-58. The fruit pericarps’ thickness is a crucial factor in determining the fruits’ firmness. Bell peppers with thick flesh may withstand lengthy shipping distances.
Table 7
Recovery of recurrent parent attributes in BC2F1 population of bell pepper for fruit traits (appraise similarity through paired t-test) Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
Pairs | Individuals | TFW (g) | FL (cm) | FW (cm) |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 757.80 | 4.52 | 3.69ns | 0.07 | 93.39 | 7.51 | 0.30 | -4.36* | 0.05 | 93.41 | 5.71 | 0.17 | 0.29ns | 0.80 | 97.72 |
LCVRS3SM-1 (BC2F1) | 707.73 | 9.34 | 8.04 | 0.27 | 5.58 | 0.28 |
2 | PAU SM-2 | 833.83 | 25.67 | 2.74ns | 0.11 | 87.94 | 7.64 | 0.15 | 4.91* | 0.04 | 91.75 | 5.92 | 0.12 | 1.62ns | 0.25 | 91.22 |
LCVRS3SM-2 (BC2F1) | 733.26 | 26.95 | 7.01 | 0.03 | 5.40 | 0.45 |
3 | PAU SM-8 | 340.50 | 3.50 | 2.92ns | 0.10 | 91.20 | 6.93 | 0.17 | 0.84ns | 0.49 | 97.11 | 3.42 | 0.00 | 2.29ns | 0.15 | 63.33 |
LCVRS3SM-8 (BC2F1) | 310.55 | 8.31 | 6.73 | 0.07 | 3.28 | 0.07 |
4 | PAU SM-4 | 540.02 | 28.22 | 0.36ns | 0.75 | 98.08 | 5.58 | 0.06 | -0.74ns | 0.54 | 93.62 | 5.68 | 0.09 | 3.54ns | 0.07 | 88.20 |
LCVRS3SM-4 (BC2F1) | 529.67 | 8.23 | 5.96 | 0.47 | 5.01 | 0.10 |
5 | PAU SM-17 | 343.33 | 29.63 | -1.59ns | 0.25 | NA | 6.82 | 0.30 | 0.17ns | 0.88 | 99.85 | 3.99 | 0.14 | -0.01ns | 0.99 | 99.75 |
LCVRS3SM-17 (BC2F1) | 370.91 | 13.96 | 6.81 | 0.23 | 4.00 | 0.32 |
6 | PAU SM-58 | 861.83 | 26.05 | 1.31ns | 0.32 | 93.40 | 6.26 | 0.13 | 0.02ns | 0.98 | 99.84 | 6.66 | 0.42 | 1.31ns | 0.32 | 87.84 |
LCVRS3SM-58 (BC2F1) | 804.91 | 30.42 | 6.25 | 0.24 | 5.85 | 0.30 |
Table 7
Pairs | Individuals | FPT | NC | FPP |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 4.13 | 0.34 | 0.74ns | 0.54 | 95.40 | 3.67 | 0.33 | 2ns | 0.18 | 81.74 | 19.33 | 0.67 | 0.65ns | 0.58 | 94.38 |
LCVRS3SM-1 (BC2F1) | 3.94 | 0.40 | 3.00 | 0.00 | 18.33 | 0.88 |
2 | PAU SM-2 | 3.75 | 0.12 | 1.56ns | 0.26 | 85.60 | 3.67 | 0.33 | 0.5ns | 0.67 | 90.74 | 14.67 | 0.67 | 0ns | 1.00 | 100 |
LCVRS3SM-2 (BC2F1) | 3.21 | 0.24 | 3.33 | 0.33 | 14.67 | 0.88 |
3 | PAU SM-8 | 3.47 | 0.06 | 0.09ns | 0.93 | 99.71 | 2.00 | 0.00 | NA | NA | 100 | 38.00 | 1.73 | 2.25ns | 0.15 | 76.32 |
LCVRS3SM-8 (BC2F1) | 3.46 | 0.12 | 2.00 | 0.00 | 29.00 | 2.65 |
4 | PAU SM-4 | 3.68 | 0.20 | 1.09ns | 0.39 | 93.75 | 3.67 | 0.33 | 0ns | 1.00 | 100 | 15.00 | 1.53 | 0.71ns | 0.55 | 88.87 |
LCVRS3SM-4 (BC2F1) | 3.45 | 0.17 | 3.67 | 0.33 | 13.33 | 0.88 |
5 | PAU SM-17 | 3.80 | 0.20 | 0.26ns | 0.82 | 98.42 | 3.33 | 0.33 | 1ns | 0.42 | 90.09 | 22.00 | 1.53 | 1.57ns | 0.26 | 83.32 |
LCVRS3SM-17 (BC2F1) | 3.74 | 0.09 | 3.00 | 0.00 | 18.33 | 0.88 |
6 | PAU SM-58 | 3.93 | 0.31 | -4.94* | 0.04 | NA | 3.67 | 0.33 | 1ns | 0.42 | 90.74 | 9.33 | 0.67 | 0ns | 1.00 | 100 |
LCVRS3SM-58 (BC2F1) | 4.70 | 0.35 | 3.33 | 0.33 | 9.33 | 0.33 |
Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
The number of lobes in the fruit of a bell pepper has an impact on the fruit's shape. In general, a fruit with four lobes, a medium size, and a blocky shape is acceptable for the market. The average number of lobes fruit− 1 for backcross populations and recurrent parents varied from 2 to 3.67 and from 2.90 to 3.50 for recurrent parents, respectively. According to per se performance, the populations LCVRS3SM-4, LCVRS3SM-58, and LCVRS3SM-2, as well as the parental lines PAU SM-1, PAU SM-2, PAU SM-4, and PAU SM-58, were found to have a higher number of lobes in the fruit. The most significant component attribute, which is directly associated to higher fruit yield plant− 1, is number of fruits plant− 1. In populations that had been backcrossed, the number of fruit plants− 1 ranged from 9.33 (LCVRS3SM-58) to 29 (LCVRS3SM-8), whereas for the parents, it ranged from 9.33 (PAUSM − 58) to 38 (PAU SM-8) (Table 7). Moreover, maximising yield per unit area is the major goal of crop cultivation in order to maximise profits. High fruit output is also the ultimate aim of any breeding programme, thus it demands the most thought. It is the primary element in farmers' adaptation to new cultivar. In case of parental lines the higher fruit yield plant− 1 was achieved by PAU SM-1 (1465.67 g) and least yield plant− 1 obtained from PAU SM-17 (764.33 g). Likewise, total fruit yield plant− 1 of the BC2F1 population was ranged from 678.18 g (LCVRS3SM-17) to 1298.35 g (LCVRS3SM-1) (Table 8).
In addition, it was observed that the trend for fruit weight, fruit width, pericarp thickness, number of locules fruit− 1, number of fruit and yield plant− 1 is similar in case of both recurrent parents and backcrossed populations which exhibited that developed populations showed no significant difference in terms of t-test analysis to their recurrent parents and it revealed that developed populations had maximum recurrent parent genome (RPG) recovery. This showed that backcrossed populations had complete phenome of their respective recurrent parents and plants of BC2F1 populations become identical to their recurrent parents for these traits. Miah et al. (2015) successfully introgressed the blast resistant Pi gene (putative Piz) into the genetic background of the widely cultivated Malaysian variety of rice (MR219), and the newly developed improved line displayed similar phenotypic traits when compared with the recurrent parent MR219, demonstrating that it was similar to the MR219 genetic backgrounds.
Table 8
Recovery of recurrent parent attributes in BC2F1 population of bell pepper for yield and total chlorophyll (appraise similarity through paired t-test)
Pairs | Individuals | Genotype Acronyms used for PCA | FYP (g) | CHT (mg 100g− 1) |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | G1 | 1465.67 | 58.77 | 1.26ns | 0.33 | 88.58 | 1.12 | 0.07 | 0.19ns | 0.87 | 99.11 |
LCVRS3SM-1 (BC2F1) | G2 | 1298.35 | 74.03 | 1.11 | 0.02 |
2 | PAU SM-2 | G3 | 1223.67 | 72.82 | 1.68ns | 0.23 | 87.50 | 0.99 | 0.03 | -13.9** | 0.01 | 81.15 |
LCVRS3SM-2 (BC2F1) | G4 | 1070.71 | 27.46 | 1.22 | 0.02 |
3 | PAU SM-8 | G5 | 905.22 | 31.77 | 0.06ns | 0.96 | 99.52 | 3.27 | 0.21 | 1.99ns | 0.18 | 89.30 |
LCVRS3SM-8 (BC2F1) | G6 | 900.85 | 85.66 | 2.92 | 0.06 |
4 | PAU SM-4 | G7 | 802.29 | 44.86 | 1.09ns | 0.39 | 88.03 | 1.57 | 0.06 | -3.06ns | 0.09 | NA |
LCVRS3SM-4 (BC2F1) | G8 | 706.25 | 47.81 | 1.80 | 0.10 |
5 | PAU SM-17 | G9 | 764.33 | 120.85 | 0.67ns | 0.57 | 88.73 | 1.69 | 0.03 | 1.99ns | 0.19 | 95.86 |
LCVRS3SM-17 (BC2F1) | G10 | 678.18 | 22.53 | 1.62 | 0.03 |
6 | PAU SM-58 | G11 | 803.80 | 59.53 | 0.8ns | 0.51 | 93.26 | 1.03 | 0.01 | -4.87* | 0.04 | NA |
LCVRS3SM-58 (BC2F1) | G12 | 749.66 | 18.16 | 1.20 | 0.05 |
Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
Continued backcrossing with the recurrent parent will be done to the significant t-test populations up until the degree of resemblance of the backcrossing progeny is very near to the recurrent parent. In order to create new LCVD resistant lines of bell pepper that are entirely isogenic to their recurrent parent in the future, the final backcross population will be self-pollinated.
Assessment of BC2F1 populations for quality attributes
A higher dry matter content correlates to a better-processed product output. Since a major amount of the chilli cultivated worldwide is consumed in powder form, fruit dry matter is a crucial feature for breeding pepper for the food processing sector (Lannes et al. 2007). The dry matter content in green fruits varied from 3.04 (PAU SM-58) to 6.04% (PAU SM-8), whereas the dry matter in red fruits ranged from 7.26 (PAU SM-58) to 14.64% (PAU SM-17) according to the parental lines mean. The similar trend for lowest and highest dry matter content in green and red fruits of the BC2F1 population was observed for LCVRS3SM-58 (4.35 & 7.57%, respectively) and LCVRS3SM-8 (6.92 & 14.82%, respectively). Furthermore, vitamin C content is also the major quality attribute in the nutritional value of bell pepper. Ascorbic acid content (mg 100g− 1) for all the six developed backcross populations ranged from 63.33 mg 100g− 1 (LCVRS3SM-17) to 74.36 mg 100g− 1 (LCVRS3SM-17), that of the recurrent parents varied from 60.19 mg 100g− 1 (LCVRS3SM-1) to 73.04 mg 100g− 1 (LCVRS3SM-17) (Table 9). The similar trend for vitamin C content in parental lines and their backcross progenies was reported by Rani (2020). While in case of capsaicin content the backcrossed populations and parental lines mean ranged from 0.2 to 0.3%, and had almost same for all the parental lines and backcross progenies. The is because the stringent negative selection was performed for capsaicin content at each backcross generation along with marker assisted selection of non-pungent plants in BC1F1 generations (Sran et al. 2023a). Hence, paired t-test analysis also dictates that there was no significance difference among the BC2F1 populations and their respective recurrent parents for capsaicin content. In context to the chlorophyll content, the highest values of chlorophyll a, b and total chlorophyll was recorded for parental line PAU SM-8 (1.11, 2.17 & 3.27 mg 100g1, respectively) and similar for its BC2F1 population (1.13, 1.79 & 2.92 mg 100g− 1, respectively). The lowest chlorophyll a and b content was noted for PAU SM-2 (0.43 & 0.99 mg 100g− 1, respectively) and LCVRS3SM-1
Table 9
Recovery of recurrent parent attributes in BC2F1 population of bell pepper for biochemical traits (appraise similarity through paired t-test) Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
Pairs | Individuals | DMG (%) | DMR (%) | VC (mg 100g− 1) |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 3.68 | 0.28 | -4ns | 0.06 | NA | 8.84 | 0.43 | -1.4ns | 0.30 | NA | 60.19 | 1.76 | -3.73ns | 0.07 | NA |
LCVRS3SM-1 (BC2F1) | 5.84 | 0.35 | 9.34 | 0.14 | 68.00 | 0.83 |
2 | PAU SM-2 | 3.45 | 0.24 | -8.1* | 0.01 | NA | 8.21 | 0.17 | -4.12ns | 0.05 | NA | 64.54 | 1.31 | -1.2ns | 0.35 | NA |
LCVRS3SM-2 (BC2F1) | 5.60 | 0.30 | 9.38 | 0.15 | 65.84 | 0.31 |
3 | PAU SM-8 | 6.04 | 0.34 | -1.18ns | 0.36 | NA | 13.69 | 0.08 | -3.45ns | 0.07 | NA | 69.72 | 1.20 | -1.84ns | 0.21 | NA |
LCVRS3SM-8 (BC2F1) | 6.92 | 0.45 | 14.82 | 0.29 | 72.08 | 0.39 |
4 | PAU SM-4 | 4.46 | 0.30 | -5* | 0.04 | NA | 11.39 | 0.24 | -1.51ns | 0.27 | NA | 62.77 | 2.40 | -1.45ns | 0.28 | NA |
LCVRS3SM-4 (BC2F1) | 5.91 | 0.11 | 11.80 | 0.31 | 65.70 | 0.89 |
5 | PAU SM-17 | 5.92 | 0.20 | -3.3ns | 0.08 | NA | 14.64 | 0.30 | 0.47ns | 0.68 | NA | 61.08 | 1.11 | -0.85ns | 0.48 | NA |
LCVRS3SM-17 (BC2F1) | 6.56 | 0.19 | 14.46 | 0.24 | 63.33 | 1.56 |
6 | PAU SM-58 | 3.04 | 0.05 | -6.13* | 0.03 | NA | 7.26 | 0.15 | -1.04ns | 0.41 | NA | 73.04 | 1.38 | -0.73ns | 0.54 | NA |
LCVRS3SM-58 (BC2F1) | 4.35 | 0.17 | 7.57 | 0.16 | 74.36 | 0.99 |
Table 9
Pairs | Individuals | CC (%) | CHA (mg 100g− 1) | CHB (mg 100g− 1) |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 0.03 | 0.00 | 0.98ns | 0.43 | 100 | 0.46 | 0.03 | 0.39ns | 0.73 | 95.65 | 0.67 | 0.05 | -0.2ns | 0.86 | 100 |
LCVRS3SM-1 (BC2F1) | 0.03 | 0.00 | 0.44 | 0.02 | 0.67 | 0.03 |
2 | PAU SM-2 | 0.04 | 0.00 | 1.24ns | 0.34 | 100 | 0.43 | 0.03 | -1.85ns | 0.21 | NA | 0.56 | 0.05 | -4.29ns | 0.05 | NA |
LCVRS3SM-2 (BC2F1) | 0.04 | 0.00 | 0.50 | 0.01 | 0.72 | 0.01 |
3 | PAU SM-8 | 0.03 | 0.00 | 1.02ns | 0.41 | 100 | 1.11 | 0.03 | -1.74ns | 0.22 | NA | 2.17 | 0.24 | 1.98ns | 0.19 | 82.49 |
LCVRS3SM-8 (BC2F1) | 0.03 | 0.00 | 1.13 | 0.02 | 1.79 | 0.08 |
4 | PAU SM-4 | 0.02 | 0.00 | -0.18ns | 0.87 | 100 | 0.85 | 0.03 | -1.97ns | 0.19 | NA | 0.72 | 0.04 | -3.05ns | 0.09 | NA |
LCVRS3SM-4 (BC2F1) | 0.02 | 0.00 | 0.94 | 0.05 | 0.86 | 0.07 |
5 | PAU SM-17 | 0.04 | 0.00 | 3.88ns | 0.06 | 75.00 | 0.72 | 0.02 | -2.33ns | 0.15 | NA | 0.97 | 0.03 | 2.2ns | 0.16 | 90.72 |
LCVRS3SM-17 (BC2F1) | 0.03 | 0.00 | 0.74 | 0.02 | 0.88 | 0.01 |
6 | PAU SM-58 | 0.04 | 0.00 | 1.67ns | 0.24 | 75.00 | 0.52 | 0.01 | -0.74ns | 0.54 | NA | 0.50 | 0.00 | -4.38* | 0.05 | NA |
LCVRS3SM-58 (BC2F1) | 0.03 | 0.00 | 0.53 | 0.01 | 0.67 | 0.04 |
Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
(0.44 & 1.11 mg 100g− 1, respectively), while lowest total chlorophyll content was recorded for parental genotype PAU SM-58 (0.50 mg 100g− 1) and BC2F1 populations LCVRS3SM-1 & LCVRS3SM-58 (0.67 mg 100g− 1) (Table 8).
The paired t-test analysis of the quality traits revealed non-significant differences for all six backcross populations with their respective recurrent parents, except for LCVRS3SM-58 for dry matter in green fruits, chlorophyll b and total chlorophyll content. Here BC2F1 population values exceed their recurrent parent mean values which suggested that parent phenome is recover during stringent phenotypic selection along with performance improvement for above said traits over the recurrent parents. It is also evident from the data presented in supplementary table 2 that fruit morphological and marketable traits had been recovered during two cycles of backcrosses. All the BC2F1 populations showed similar fruit and morphological characteristics as their respective recurrent parents. Fruit characteristics like fruit shape and color is vital attributes for market acceptability in case of bell pepper hence it is important to recover these traits along with the gene of interest. In the studies of the gene introgression from hot pepper to bell pepper the linkage drag for fruit traits is the most major drawback of hot pepper and bell pepper hybridization programmes (Lapidot et al. 1997). Hence the stringent phenotypic selection for these traits proved to be beneficial in hot pepper and bell pepper breeding programmes.
Assessment of BC2F1 populations for seed attributes
Besides from LCVD resistance, the good quality seed production is also an important aspect from the breeder and seed industry point of view, especially when we breed the bell pepper genotype for heat tolerance. Hence the numbers of seeds fruit− 1, 1000 seed fresh and dry weight are important traits that were evaluated. The number of seed fruit− 1 ranged from 55 (PAU SM-8) to 78.33 (PAU SM-4) in parental lines and among the BC2F1 populations the same trend was observed and the highest number of seed fruit− 1 was witnessed for LCVRS3SM-4 (77.67) and lowest was noted for LCVRS3SM-8 (53.33). With the exception of one population (LCVRS3SM-8), the mean values of developed backcrossed populations carrying the LCVD resistance gene showed statistically non-significant differences to their recurrent parents. This indicated that the developed progenies have the
Table 10
Recovery of recurrent parent attributes in BC2F1 population of bell pepper for seed traits (appraise similarity through paired t-test)
Pairs | Individuals | SPF | SFW (g) | SDW (g) |
Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) | Mean | SEm± | t | p-value | RPP-R (%) |
1 | PAU SM-1 | 66.00 | 3.21 | 0.76ns | 0.53 | 96.97 | 13.66 | 0.21 | 10.87** | 0.01 | 93.05 | 8.67 | 0.19 | 2.22ns | 0.16 | 96.08 |
LCVRS3SM-1 (BC2F1) | 64.00 | 1.15 | 12.71 | 0.13 | 8.33 | 0.05 |
2 | PAU SM-2 | 60.67 | 2.33 | 1.26ns | 0.33 | 93.95 | 10.79 | 0.29 | 0.08ns | 0.95 | 99.63 | 7.67 | 0.18 | 2.3ns | 0.15 | 88.79 |
LCVRS3SM-2 (BC2F1) | 57.00 | 3.21 | 10.75 | 0.14 | 6.81 | 0.20 |
3 | PAU SM-8 | 55.00 | 2.08 | 5* | 0.04 | 96.49 | 6.69 | 0.04 | 1.43ns | 0.29 | 89.99 | 5.52 | 0.03 | 9.89* | 0.01 | 78.62 |
LCVRS3SM-8 (BC2F1) | 53.33 | 2.19 | 6.02 | 0.45 | 4.34 | 0.10 |
4 | PAU SM-4 | 78.33 | 2.40 | 0.16ns | 0.89 | 99.16 | 7.36 | 0.11 | -2.21ns | 0.16 | NA | 6.41 | 0.19 | 1.15ns | 0.37 | 97.04 |
LCVRS3SM-4 (BC2F1) | 77.67 | 1.86 | 7.73 | 0.25 | 6.22 | 0.03 |
5 | PAU SM-17 | 63.67 | 2.73 | -0.34ns | 0.77 | NA | 8.54 | 0.08 | 0.87ns | 0.48 | 97.66 | 5.83 | 0.06 | 2ns | 0.18 | 94.00 |
LCVRS3SM-17 (BC2F1) | 65.67 | 3.18 | 8.34 | 0.17 | 5.48 | 0.12 |
6 | PAU SM-58 | 63.33 | 2.40 | -0.25ns | 0.83 | NA | 11.89 | 0.09 | 6.28* | 0.02 | 92.09 | 9.35 | 0.04 | 14.94** | 0.00 | 85.13 |
LCVRS3SM-58 (BC2F1) | 64.33 | 2.85 | 10.95 | 0.08 | 7.96 | 0.06 |
Note: *Significant at the 5% level of significance (p < 0.05); ns: non-significant at 5% level of significance; SEm±: Standard error of the mean; RPP_R: recurrent parent phenome recovery; For traits acronyms see Table 2
same good seed setting ability as their parental lines of bell pepper and also recover the genome after two backcrosses. Secondly, the test weight of fresh and dry seed for parental lines was ranged from 6.69 g (PAU SM-8) to 13.66 g (PAU SM-1) and 5.52 g (PAU SM-8) to 9.35 g (PAU SM-58), respectively. Likewise, the similar trend of highest and lowest range for fresh and dry test weight of seed was observed in BC2F1 populations (Table 10). Based on t-test analysis, it was recorded that the developed BC2F1 populations had similar fresh and dry seed test weight except for LCVRS3SM-1 for fresh seed test weight, LCVRS3SM-8 for dry seed test weight and LCVRS3SM-58 for both. Consequently, backcrossing will continue with their respective recurrent parents until the degree of resemblance between the backcrossing progeny and the recurrent parent is achieved.
Evaluation of fruit traits across the generations
To determine the trend of recovery of the recurrent parent phenome (RPP), the morphological traits of the donor parent (DP), hot pepper line S-343, and the recurrent parent (RP), bell pepper lines PAU SM-1, PAU SM-2, PAU SM-8, PAU SM-4, PAU SM-17, and PAU SM-58, were evaluated. The change in means for plant growth and fruit attributes was used to determine the recovery rate of the recurrent parent in BC1F1 and BC2F1 progenies. Figure 2 displays the mean trait values of the parent, F1, BC1F1, and BC2F1 populations with an LSD 5% analysis. In case of all the studied fruit traits the performance of BC2F1 was comparatively near to the respective recurrent parents. The ten fruit weight was found to be higher in PAU SM-58 (861.83 g) and minimum in donor parent S-343 (43.67 g) (Supplementary Table 2). The performance of F1, BC1 F1 and BC2 F1 generations for ten fruit weight is 226.17 g, 380.75 g and 576.17 g, respectively. While in case of fruit length values of backcross populations were close to both the parents. The highest fruit length was observed for F1 generations (9.61 cm). On the other hand, for fruit width the BC2F1 generations general mean (4.85 cm) is almost similar to the parental lines general mean (4.68 cm), indicated the complete recovery of the fruit shape in advanced backcross populations. The lowest mean value for fruit width was observed for the F1 generations (2.51 cm). This is quite evident from the figures of fruits and plants of hot pepper and bell pepper parents along with F1, BC1F1, and BC2F1 generations published in the work done by the Sran and co-workers (2023a). Likewise, for fruit pericarp thickness comparative performance of BC2F1
Figure 2 (a-g) Comparative performance of recurrent parents, F1, BC1F1 and BC2F1 generations
progenies (3.75 mm) was more similar to recurrent parents’ average value (3.79 mm) than donor parent (1.18 mm) as compare to F1 hybrid (2.05 mm) and BC1F1 backcross (2.56 mm). The perusal of mean values for number of locules fruit− 1 of donor parent (S-343), recurrent parents, F1 crosses, BC1F1 and BC2F1 generations were 2, 3.33, 2.28, 2.56 and 3.06, respectively (Supplementary Table 2). Furthermore, the number of fruit plant− 1 was recorded higher in donor parent (S-343) as compared to recurrent bell pepper parents, also the average value of F1 for the number of fruits plant− 1 was found to be higher (24.28) than BC1F1 (11.89) and BC2F1 (12.36) generations. Finally, for the most important trait the fruit yield plant− 1 the recurrent parent phenome (RPP) was recover upto 90.59% in BC2F1 generations with mean yield plant− 1 of 900.67 g, while for F1 and BC1F1 the RPP recovery was 57.81% and 45.68%, respectively. The RPP recovery was reported to be up to 91.6% in BC2F1 generation for pyramiding blast resistance genes Piz5 and Pi54 into an elite Basmati rice restorer line "PRR78" in various investigations carried out by other research groups (Singh et al. 2013). Our outcomes concurred with those of Ahmed et al. (2016), who obtained recovery rates for BC1F1 and BC2F1 populations of 65.55 to 77.8 and 78.79 to 95.5%, respectively. In another research work done by Meena and co-workers (2019) they achieved the recovery of 96.6 to 98.8% of recurrent parent genome in BC5F1 generations during introgression of male sterility gene from hot pepper to bell pepper genotypes.
Correlation analysis
The understanding of correlations among various attributes in parental lines and backcross populations is just as crucial as the recovery of the recurring parent phenome, and genetic diversity among these. Figure 3 shows the correlation graph for the growth, yield, fruits and seed attributes using Pearson's correlation coefficient (P < 0.05), and Supplementary Table 3 contains the results. The correlation analysis revealed a significant positive relation of plant growth traits like plant height (PH) with number of fruits plant− 1, dry matter green and red fruits and significant negative correlation with FW, VC and SDW. Other plant growth trait like PW depicted significant correlation with FPP, CHB and CHT, while NB showed positive correlation with CHA and negative correlation with CC. Among fruit traits the positive significant correlation was seen between FW, TFW, NL, SFW and SDW while FW and TFW showed significant negative relation with PH, FPP, DMG, DMR, CHA, CHB and CHT. Correlation analysis revealed that increase in plant height and spread also increases the number of fruits palnt− 1 while fruit weight was directly influenced by fruit width, number of locules fruit− 1, 1000 seed fresh and dry weight. Among biochemical attributes, significant positive correlation was noted between DMG, DMR, CHA, CHB and CHT however these traits depicted negative relation with FW, TFW, NL, SFW and SFW. The similar findings for biochemical attributes were also reported by Sran
and Jindal (2022). On the other hand, the 1000 seed dry and fresh weight was positively influenced by fruit width and ten fruit weight and negatively influenced by dry matter content in green and red fruits and chlorophyll content.
Cluster analysis and PCA
Cluster analysis was performed to examine the variability present among the developed backcross populations and their respective recurrent parents and to cluster their growth, fruit, quality, and seed attribute data recorded during stringent phenotypic selection for recurrent parent’s phenome. The 12 genotypes including both recurrent parents and their respective BC2F1 progenies were divided into two major classes and into four distinct groups. Group A contains 10 genotypes while group B have 2 genotypes (Fig. 4). It is interesting to observe that hierarchical cluster analysis grouped each BC2F1 generation with its respective recurrent parent and so it is evident from the cluster analysis of recorded data of 20 growth and quality traits that the recurrent parent phenome was recovered through stringent phenotypic selection during the backcrossing with recurrent parents. Cluster A subdivided the genotypes into three groups on the basis of fruit traits, which are rectangular and squared-fruit-shaped genotypes while cluster B consisted of genotypes with cordate-fruit-shaped. Moreover, these recorded traits are grouped into two major clusters that are subdivided into three distinct groups (Fig. 4). The fruit and seed traits are grouped in cluster A while biochemical and plant growth parameters grouped into cluster B. The fruit length and fruit yield plant− 1 clustered together and dry matter content in green and red fruit grouped with plant height, similarly chlorophyll content and fruits plant− 1 grouped together. The correlation analysis has also revealed significant positive relations among these clustered traits. Hence, it can be affirmed that grouping of parents and their progenies can be done on the basis of recorded yield, growth, fruit, quality and seed attributes. Therefore, it is conclusive to perform selection for fruit and quality traits for the recovery of phenome of recurrent parents.
The experiment's principal component is a multivariate statistical technique for examining intricate and substantial data sets. Twelve distinct genotypes and progenies were assigned to twenty set of characteristics, producing vast amounts of data. In order to determine the link between the several variables that create variability in the tested data, PCA was also used to investigate the variation pattern among the tested genotypes and characteristics. Over 97% of the variance in the first six components was explained by the PCA (Supplementary table 4, Fig. 5A) yet 100% variation was explained by total twelve PCA components. The first two PC components (PC1 & PC2) in Fig. 5A for both genotypes and studied traits showed 69.33% variation while rest of the components explained 30.67% variation. Among the tested growth and quality attributes CHA, FW, FPP, SFW, CHT, CHB, SDW, TFW, DMR, NL, FL and SPF were the major contributing traits to the total observed variability in PC1 and PC2 (Supplementary Fig. 1 ,and Fig. 5B). While, in case of tested parental lines and backcross progenies G3 and its BC2F1 progeny G9 followed by G6, G4 and its BC2F1 progeny G10 were the major genotypes that contributing to the observed variability (Supplementary Fig. 2 ,and Fig. 5B). The treatments and attributes were represented as vectors in 2-D biplots, and their relative distance from the origin indicated how much of the observed variability they contributed. It is interesting to observed that the parental lines and their respective recurrent progenies lies closer to each other on biplots and attributes that were clustered together by cluster analysis such as PW, CHA, FPP, CHT, CHB, DMG & DMR, and FPT, SDW & SFW were also lied close to each other toward
Figure 5B Eigenvalues and cumulative variability for all twelve principal components of principal component analysis. (For genotypes acronyms see Table 8 & for traits acronyms see Table 2)
Figure 5A Biplot based on PC1 and PC2 represents variation of different studied traits and genotypes, respectively.
same direction on PC1 & PC2 components biplot and respectively, exhibited their correlation among themselves. The current study's principal component analysis (PCA) results were consistent with those of previous research using PCA to examine genotype and trait variability in pepper and other crops (Zafer et al. 2021; Sran and Jindal 2022; Sran et al. 2023b).